There has long been a need for satisfactory methods of disrupting viruses, especially lipid-containing viruses, for example, the viruses of influenza, Newcastle disease, rabies, and the like. Available methods for the production of virus antigens involve isolation of virus from natural sources or following propagation of virus in fowl embryo cell fluids, tissue culture fluids and similar aqueous media.
The viruses formed in these media consist of discrete particles, which contain, for example, bound-in lipids. More recently, improved vaccines have been prepared by disrupting the intact viruses into sub-units, which are still desirably antigenic.
Several means are now used on large scale to achieve the desired disruption of viruses into sub-units. These comprise generally the use, on the one hand, of detergents or, on the other, ether, a volatile highly-flammable substance. A detergent-disrupted, sub-unit vaccine prepared by sodium desoxycholate treatment, of influenza viruses is commercially available but suffers from considerably reduced immunogenicity (as measured by mouse potency tests) as compared with the intact viruses (Rubin et al., Arch. Virus-forsch., 20:268 [1967]; Webster, J. Immunol., 96:596 [1966]). Another important proposal has been to use ether as a disrupting agent. However, contrary to some claims in the literature, and as will be shown in the drawing, ether does not nearly completely eliminate toxicity as measured by standard pyrogenicity tests in rabbits. Furthermore, ether can cause a substantial loss of the enzyme, neuraminidase. This is disadvantageous because it has been suggested that the best vaccines must contain good levels of this enzyme. Therefore, it would be desirable to provide means to disrupt viruses, especially influenza viruses, into sub-units, which means are more effective than ether for removing pyrogens, without at the same time causing substantial losses of neuraminidase, and which do not reduce the potency of the virus as an immunizing agent, as do detergents such as sodium desoxycholate. It is also desirable to provide means to disrupt concentrated viruses as well as impure allantoic fluids. Surprisingly, such means for the disaggregation of viruses are provided by the method of the instant invention.
It is, therefore, a primary object of the instant invention to provide methods to disrupt viruses.
It is another object of the instant invention to provide a means to disrupt viruses at the same time lowering toxicity and pyrogenicity.
Still another object of the instant invention is to provide a means to disrupt viruses without at the same time lowering significantly the neuraminidase level.
It is a further object of the instant invention to provide methods to disrupt viruses without substantially decreasing the immunogenicity thereof as compared with intact viruses.
Still another object of the instant invention is to provide a means to disrupt viruses which can be employed in large scale manufacturing facilities without danger from explosion or fire.
Still another object of the instant invention is to provide disrupted viruses useful to prepare vaccines for diagnostic and immunogenic purposes.